Benzo [a] pyrene is a potent mutagen/carcinogen and reacts with DNA via its (+)-anti-7, 8-diol-9, 10-epoxide ( (+)-anti-B{a}PDE) to give DNA adducts, principally at N2 -Gua and N6 -Ade. The mechanisms of mutagenesis by (+)-anti-B{a}PDE was investigated using both site-specific and random mutagenesis approaches. (10 An E. coli plasmid was constructed that contained (+)-anti-B{a}P-N2-Gua in a 5'-TGC-3' sequence context and was used to determine the G->T mutations were virtually exclusively induced. (2) random mutagenesis by (+)-anti-B{a}PDE was studied in a system we developed, where a plasmid a supF target gene was adducted in vitro and transformed into E. coli. G->T mutations predominated in 5'-TG-3"""""""" sequence contexts, which correlated with the site-specific studies and suggested that (+)-anti-B{a}P-N2-Gua was responsible. In contrast, 5'-GG-3', 5'-CG-3' and 5'-AG-3' sequence contexts showed a significant fraction of GC->AT and GC->CG mutations. G115 was the major base pairing hot-spot. Some mutational complexity suggested either: (1) that there are two possible conformations for an adduct at G115 with different mutational outcomes; or (2) that there is a labile adduct at G115, which is converted to an AP-site. Preliminary data suggests that the former is more likely. (+)-anti-B{a}PDE generates labile adducts, which are probably not N7-Gua adducts. It is more likely that N2-Gua adducts can adopt multiple conformations, where one is stable, while the other is labile. This may relate to the evidence for multiple conformations obtained in mutagenesis studies at G115. The following unifying hypothesis will be tested. A single adduct can adopt multiple conformations, each of which can cause a different kind of mutation (s). Furthermore, adduct conformation can be influenced by DNA sequence context, as well as environmental factors, such as heating.

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Sholder, Gabriel; Creech, Amanda; Loechler, Edward L (2015) How Y-Family DNA polymerase IV is more accurate than Dpo4 at dCTP insertion opposite an N2-dG adduct of benzo[a]pyrene. DNA Repair (Amst) 35:144-53
Sholder, Gabriel; Loechler, Edward L (2015) A method to accurately quantitate intensities of (32)P-DNA bands when multiple bands appear in a single lane of a gel is used to study dNTP insertion opposite a benzo[a]pyrene-dG adduct by Sulfolobus DNA polymerases Dpo4 and Dbh. DNA Repair (Amst) 25:97-103
Ikeda, Mio; Furukohri, Asako; Philippin, Gaelle et al. (2014) DNA polymerase IV mediates efficient and quick recovery of replication forks stalled at N2-dG adducts. Nucleic Acids Res 42:8461-72
Chandani, Sushil; Loechler, Edward L (2013) Structural model of the Y-Family DNA polymerase V/RecA mutasome. J Mol Graph Model 39:133-44
Seo, Kwang Young; Yin, Jun; Donthamsetti, Prashant et al. (2009) Amino acid architecture that influences dNTP insertion efficiency in Y-family DNA polymerase V of E. coli. J Mol Biol 392:270-82
Chandani, Sushil; Loechler, Edward L (2009) Y-Family DNA polymerases may use two different dNTP shapes for insertion: a hypothesis and its implications. J Mol Graph Model 27:759-69
Clapp, Richard W; Jacobs, Molly M; Loechler, Edward L (2008) Environmental and occupational causes of cancer: new evidence 2005-2007. Rev Environ Health 23:1-37
Herscovitch, Melanie; Comb, William; Ennis, Thomas et al. (2008) Intermolecular disulfide bond formation in the NEMO dimer requires Cys54 and Cys347. Biochem Biophys Res Commun 367:103-8
Chandani, Sushil; Loechler, Edward L (2007) Molecular modeling benzo[a]pyrene N2-dG adducts in the two overlapping active sites of the Y-family DNA polymerase Dpo4. J Mol Graph Model 25:658-70
Kalam, M Abul; Haraguchi, Kazuhiro; Chandani, Sushil et al. (2006) Genetic effects of oxidative DNA damages: comparative mutagenesis of the imidazole ring-opened formamidopyrimidines (Fapy lesions) and 8-oxo-purines in simian kidney cells. Nucleic Acids Res 34:2305-15

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